# Proton channels govern vesicular carbonate chemistry in mineralizing cells of a marine calcifier

## TL;DR

This study reveals how proton channels control the chemistry inside cells that form calcium carbonate in sea urchin embryos.

## Contribution

The discovery of Otop2l as a proton channel regulating alkaline pH in calcifying vesicles is novel in biomineralization research.

## Key findings

- Calcifying cells and vesicles in sea urchin embryos show high proton permeability.
- Otop2l is identified as a proton channel activated by alkaline pH and Ca2+/Mg2+ ions.
- High proton conductance helps maintain alkaline conditions in calcification vesicles.

## Abstract

Biomineralization shapes the geology of our planet and is an integral part of the global carbon cycle. Many calcifying organisms generate biomineral precursors within their cells but the mechanisms controlling carbonate chemistry in the calcifying vesicles remain unknown. Using the sea urchin embryo that is a prime model for biomineralization by intracellular amorphous calcium carbonate (ACC) formation this work demonstrated a high proton permeability of the calcifying cells and their vesicles. Intra-vesicular pH and Ca2+ recordings demonstrate highly alkaline conditions in ACC forming endocytotic Ca2+-rich vesicles, that are enriched during the mineralization process. Using confocal live-cell imaging we observe how vesicles exocytose their Ca2+-rich and alkaline contents to the calcification front. We identify the proton channel Otop2l as a master regulator for membrane proton conductance in calcifying cells, using the membrane potential to control pH conditions at the site of mineral precursor formation. Electrophysiological investigations demonstrate that Otop2l is activated by alkaline conditions as well as Mg2+ and Ca2+ ions. We provide evidence for high intracellular proton conductance as mechanism to generate alkaline pH conditions in the calcification vesicles. This deep mechanistic knowledge can help explain sensitivities of marine calcifiers to rapid changes in seawater pH in past and future marine habitats.

Tight regulation of carbonate chemistry is required for biomineralization to occur. Here, the authors identify proton channel as key regulator of high intracellular proton conductance and maintenance of alkaline pH in the calcifying vesicles of larval sea urchin cells.